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PubMed1995

Construction of species-specific primers for Pseudomonas andropogonis based on 16S rDNA sequences.

Approximately 94% of the total 16S rDNA of Pseudomonas andropogonis strain ACH 01053A was sequenced and compared with that of strain ATCC 23061 obtained from the GenBank database. The two sequences were highly homologous with 1.3% difference. Alignment of sequences with those from closely related bacteria revealed two possible regions for design of a specific primer suitable for detection of Ps. andropogonis. Using primers designed to these variable regions in a PCR test, an amplification product of approximately 410 bp was specifically produced by 40 strains of Ps. andropogonis. No other bacterial species showed an amplification product under optimized PCR conditions. As few as 1000 cells per reaction were detected.

Base Sequence↗

PubMed1993

Phylogenetic diversity of the genus Cytophaga revealed by 16S rRNA sequencing and menaquinone analysis.

To clarify the intra- and intergeneric relationships of the genus Cytophaga, 16S rRNA sequences and respiratory isoprenoid quinones were determined for the type strains of the 21 validly published species and one isolate in the genus Cytophaga. The sequence analysis revealed extreme heterogeneity of this genus, which diverged into nine distinct lines of descent. Each lineage of Cytophaga was characterized by possessing either menaquinone-6 (MK-6) or MK-7. The MK-6-possessing species were located in the two lineages that were remote from MK-7 species. One of the MK-6 lineages was composed only of terrestrial species and the other only of marine species. Flavobacterium aquatile, the type species of the genus Flavobacterium, was located in the MK-6 terrestrial lineage. The terrestrial Cytophaga species with MK-6 should be transferred to the genus Flavobacterium. The marine facultative anaerobes with MK-7 were located in the bacteroides branch, and possessed signature sequences with features intermediate between the bacteroides and the flavobacteria subdivisions. Cytophaga hutchinsonii, the type species of the genus Cytophaga, had a close relationship only with Cytophaga aurantiaca. The genus Cytophaga should be restricted to these two cellulose-degrading species. The genus Cytophaga is so heterogeneous that it should be divided into several genera and higher taxa in accordance with the phylogenetic relationships.

Base Sequence↗

PubMed1994

A phylogenetic tree of 16S rRNA sequences from sulfate-reducing bacteria in a sandy marine sediment.

The divergence of 16S rDNA sequences in marine sediment was investigated. Twenty unique partial sequences were found among 33 cloned following PCR. Thirteen shared 82 to 91% similarity with sequences of delta subclass sulfate-reducing bacteria. Three contained the target sequence for a sulfate-reducing bacterium-specific oligonucleotide probe designed from pure-culture studies.

Bacteria↗

PubMed1998

Phylogeny of the family Moraxellaceae by 16S rDNA sequence analysis, with special emphasis on differentiation of Moraxella species.

Thirty-three strains previously classified into 11 species in the bacterial family Moraxellaceae were subjected to phylogenetic analysis based on 16S rRNA sequences. The family Moraxellaceae formed a distinct clade consisting of four phylogenetic groups as judged from branch lengths, bootstrap values and signature nucleotides. Group I contained the classical moraxellae and strains of the coccal moraxellae, previously known as Branhamella, with 16S rRNA similarity of > or = 95%. A further division of group I into five tentative clusters is discussed. Group II consisted of two strains representing Moraxella atlantae and Moraxella osloensis. These strains were only distantly related to each other (93.4%) and also to the other members of the Moraxellaceae (< or = 93%). Therefore, reasons for reclassification of these species into separate and new genera are discussed. Group III harboured strains of the genus Psychrobacter and strain 752/52 of [Moraxella] phenylpyruvica. This strain of [M.] phenylpyruvica formed an early branch from the group III line of descent. Interestingly, a distant relationship was found between Psychrobacter phenylpyruvicus strain ATCC 23333T (formerly classified as [M.] phenylpyruvica) and [M.] phenylpyruvica strain 752/52, exhibiting less than 96% nucleotide similarity between their 16S rRNA sequences. The establishment of a new genus for [M.] phenylpyruvica strain 752/52 is therefore suggested. Group IV contained only two strains of the genus Acinetobacter. Strategies for the development of diagnostic probes and distinctive sequences for 16S rRNA-based species-specific assays within group I are suggested. Although these findings add to the classificatory placements within the Moraxellaceae, analysis of a more comprehensive selection of strains is still needed to obtain a complete classification system within this family.

DNA Primers↗

PubMed2001

Specific 16S rDNA sequences associated with naphthalene degradation under sulfate-reducing conditions in harbor sediments.

Previous studies have demonstrated that naphthalene and other polycyclic aromatic hydrocarbons (PAHs) can be anaerobically oxidized with the reduction of sulfate in PAH-contaminated marine harbor sediments, including those in San Diego Bay. In order to learn more about the microorganisms that might be involved in anaerobic naphthalene degradation, the microorganisms associated with naphthalene degradation in San Diego Bay sediments were evaluated. A dilution-to-extinction enrichment culture strategy, designed to recover the most numerous culturable naphthalene-degrading sulfate reducers, resulted in the enrichment of microorganisms with 16S rDNA sequences in the d-Proteobacteria, which were closely related to a previously described pure culture of a naphthalene-degrading sulfate reducer, NaphS2, isolated from sediments in Germany. A more traditional enrichment culture approach, expected to enrich for the fastest-growing naphthalene-degrading sulfate reducers, yielded 16S rDNA sequences closely related to those found in the dilution-to-extinction enrichments and NaphS2. Analysis of 16S rDNA sequences in sediments from two sites in San Diego Bay that had been adapted for rapid naphthalene degradation by continual amendment with low levels of naphthalene suggested that the microbial community composition in the amended sediments differed from that present in the unamended sediments from the same sites. Most significantly, 6-8% of the sequences recovered from 100 clones of each of the naphthalene-amended sediments were closely related to the 16S rDNA sequences in the enrichment cultures as well as the sequence of the pure culture, NaphS2. No sequences in this NaphS2 phylotype were recovered from the sediments that were not continually exposed to naphthalene. A PCR primer, which was designed based on these phylotype sequences, was used to amplify additional 16S rDNA sequences belonging to the NaphS2 phylotype from PAH-degrading sediments from Island End River (Boston), MA, and Liepaja Harbor, Latvia. Closely related sequences were also recovered from highly contaminated sediment from Tampa Bay, FL. These results suggest that microorganisms closely related to NaphS2 might be involved in naphthalene degradation in harbor sediments. This finding contrasts with the frequent observation that the environmentally relevant microorganisms cannot be readily recovered in pure culture and suggests that further study of the physiology of NaphS2 may provide insights into factors controlling the rate and extent of naphthalene degradation in marine harbor sediments.

Biodegradation, Environmental↗

PubMed2005

[Bacterial 16S rDNA sequence analysis of Siberian tiger faecal flora].

Bacterial 16S rDNA library of Siberian tiger was developed and 15 different clones were obtained using EcoR I and Hind III in restriction fragment length polymorphism analysis. DNA sequencing and similarity analysis showed that 10 clones matched corresponding Clostridium sequences, of which 6 sequences had over 99% similarity with Clostridium novyi type A, and 4 sequences had 97% similarity with Swine manure bacterium RT-18B, which identified as Peptostreptococcus spp. The other five 16S rDNA sequences had 94% - 95% similarity with Clostridium pascui, Clostridium tetani E88, Clostridium sp. 14505 Clostridium perfringens and Carnobacterium sp. R-7279 respectively.

Animals↗

PubMed2002

Cloning and sequencing of 16S rDNA and 16S-23S rDNA internal spacer region (ISR) from urease-positive thermophilic Campylobacter (UPTC).

AIMS: To clone and sequence the 16S rDNA and 16S-23S rDNA internal spacer region (ISR) from urease-positive thermophilic Campylobacter (UPTC). METHODS AND RESULTS: The primer sets for 16S rDNA and 16S-23S rDNA ISR amplified almost the full length of 16S rDNA and 16S-23S rDNA ISR. About 1500 bp for 16S rDNA and about 720 bp for 16S-23S rDNA ISR of the rrn operon of four strains of UPTC were identified after molecular cloning and sequencing. CONCLUSIONS: The four strains and CCUG18267 of UPTC showed approximately 99% sequence homology of 16S rDNA to each other, 96-97% to Camp. coli, 97-98% to Camp. jejuni and 97-98% to Camp. lari. SIGNIFICANCE AND IMPACT OF THE STUDY: For the first time, the nucleotide sequence of 16S-23S rDNA ISR of UPTC has been analysed. The sequence of ISR was almost identical among the four strains of UPTC. It is interesting that the UPTC intercistronic tRNAs demonstrated an order of tRNA of 5'-16S-tRNAAla-tRNAIle-23S-3' in the organisms.

Campylobacter↗

PubMed2004

Identification of three clusters of canine intestinal spirochaetes by biochemical and 16S rDNA sequence analysis.

It has been suggested that canine intestinal spirochaetes consist of Brachyspira pilosicoli and a group of strains that has been provisionally designated 'Brachyspira canis'. The purpose of the present study was to compare 22 spirochaete isolates that were obtained from intestinal specimens of dogs in Sweden (n = 12), Norway (n = 4), the United States (n = 3), Australia (n = 2) and Germany (n = 1) with type and reference strains, as well as field isolates, of Brachyspira species by five biochemical tests and determination of almost-complete 16S rDNA sequences. In an evolutionary tree derived from 16S rDNA sequences, the canine isolates grouped into three clusters. One cluster included the type strain of porcine B. pilosicoli, whereas a second larger cluster, which was monophyletic, contained a canine strain that was identified previously as 'B. canis'. The third cluster consisted of three canine isolates of Scandinavian origin, which grouped together with the type strain of the species Brachyspira alvinipulli (pathogenic to chicken). These three genotypes, which were identified on the basis of 16S rDNA sequences, corresponded to four phenotypic groups based on biochemical testing. Two biochemical tests, hippurate hydrolysis and alpha-galactosidase production, were sufficient for rapid identification of each canine cluster.

Animals↗

PubMed1985

The sequence of 16S rRNA from Mycoplasma strain PG50.

The DNA sequence of one of the 16S rRNA genes (cistron rrnA) of Mycoplasma strain PG50 was determined. It is 1523 bp long and has about 70% homology to the sequence of Escherichia coli 16S rRNA (rrnB). The G + C content of the sequence is 48% compared with 56% G + C in the E. coli sequence. The secondary structure is formed and it is determined that most of the differences between the two sequences are seen in stems while loops in the secondary structure are conserved. A detailed description of differences and similarities to known sequences and rRNA oligonucleotide catalogues is given, and this information is used to discuss functional properties and phylogenetic relations of mycoplasma 16S rRNA.

Base Sequence↗

PubMed1992

How close is close: 16S rRNA sequence identity may not be sufficient to guarantee species identity.

16S rRNA (genes coding for rRNA) sequence comparisons were conducted with the following three psychrophilic strains: Bacillus globisporus W25T (T = type strain) and Bacillus psychrophilus W16AT, and W5. These strains exhibited more than 99.5% sequence identity and within experimental uncertainty could be regarded as identical. Their close taxonomic relationship was further documented by phenotypic similarities. In contrast, previously published DNA-DNA hybridization results have convincingly established that these strains do not belong to the same species if current standards are used. These results emphasize the important point that effective identity of 16S rRNA sequences is not necessarily a sufficient criterion to guarantee species identity. Thus, although 16S rRNA sequences can be used routinely to distinguish and establish relationships between genera and well-resolved species, very recently diverged species may not be recognizable.

Bacillus↗

PubMed2000

[Reconstruction of phylogeny of locusts from the family Acrididae (Orthoptera) based on analysis of nucleotide sequences of 16S ribosome RNA gene in mitochondria].

The sequences of two mitochondrial 16S RNA gene fragments (137- and 174-bp in size) were determined in nine grasshopper species belonging to three Acrididae subfamilies. Phylogenetic reconstruction was performed using the sequences of twelve grasshopper species and the cricket Acheta domesticus sequence as an outgroup (some data were purchased from the GeneBank Data Library (NCBI). In the phylogenetic tree, the Acridinae and Locustinae formed compact groups. Annexpected position of Celes scalozubovi (Locustinae) within the subfamily Acridinae indicated its vague phylogeny. The Catantopinae species lied close to the base of the Acridinae. Almost all branches of phylogenetic trees were strongly (55-100%) supported by bootstrap analysis.

Animals↗

PubMed1995

Characterization of chemoautotrophic bacterial symbionts in a gutless marine worm Oligochaeta, Annelida) by phylogenetic 16S rRNA sequence analysis and in situ hybridization.

The phylogenetic relationships of chemoautotrophic endosymbionts in the gutless marine oligochaete Inanidrilus leukodermatus to chemoautotrophic ecto- and endosymbionts from other host phyla and to free-living bacteria were determined by comparative 16S rRNA sequence analysis. Fluorescent in situ hybridization confirmed that the 16S rRNA sequence obtained from these worms originated from the symbionts. The symbiont sequence is unique to I. leukodermatus. In phylogenetic trees inferred by both distance and parsimony methods, the oligochaete symbiont is peripherally associated with one of two clusters of chemoautotrophic symbionts that belong to the gamma subdivision of the Proteobacteria. The endosymbionts of this oligochaete form a monophyletic group with chemoautotrophic ectosymbionts of a marine nematode. The oligochaete and nematode symbionts are very closely related, although their hosts belong to separate, unrelated animal phyla. Thus, cospeciation between the nematode and oligochaete hosts and their symbionts could not have occurred. Instead, the similar geographic locations and habitats of the hosts may have influenced the establishment of these symbioses.

Animals↗

PubMed1994

Evolutionary affiliation of the marine nitrogen-fixing cyanobacterium Trichodesmium sp. strain NIBB 1067, derived by 16S ribosomal RNA sequence analysis.

The 16S rRNA sequence of Trichodesmium sp. strain NIBB 1067 was determined and used for the construction of a distance tree and bootstrap analysis. The tree shows that, among the available cyanobacterial 16S rRNA sequences, Trichodesmium NIBB 1067 has Oscillatoria PCC 7515 as its closest relative, presenting 94.9% of sequence similarity with the latter strain. This is in contrast to a difference of 9 mol% G+C in mean genomic DNA base composition between the two organisms. Nevertheless, the genotypic heterogeneity presented by a number of strains assigned to the genus Oscillatoria hinders a taxonomic decision on the separate existence of the genera Trichodesmium and Oscillatoria. The sequence of the internal transcribed spacer (ITS) between the 16S and 23S rRNA genes was also determined, as a possible marker to study inter- and intraspecific variability. The ITS contains the genes coding for tRNA(Ile) and tRNA(Ala) and its total length is 547 nucleotides. In six out of eight sequenced clones, there is a duplication of 29 nucleotides, surrounding the 5' end of the tRNA(Ile).

Base Sequence↗

PubMed1991

Oligodeoxynucleotide probes for Campylobacter fetus and Campylobacter hyointestinalis based on 16S rRNA sequences.

Deoxyoligonucleotide probes were constructed for the identification of Campylobacter fetus and Campylobacter hyointestinalis based on 16S rRNA sequence data. Probes were targeted to hypervariable regions of 16S rRNA. Specificity of oligonucleotide probes was tested in a colony blot assay with type strains of 15 Campylobacter and Arcobacter species as well as in a slot blot format using genomic DNA extracted from field strains of C. fetus and C. hyointestinalis. Two oligonucleotides were constructed for C. fetus that hybridized with equal specificity with each of 57 biochemically confirmed isolates of C. fetus but not with any other Campylobacter species. The C. hyointestinalis probe reacted with 47 of 48 biochemically confirmed field isolates of C. hyointestinalis. In Southern blot hybridization of BglII digests of genomic DNA, the respective probes reacted within three restriction fragments of either C. hyointestinalis (7.2, 8.2, and 10.1 kb) or C. fetus (7.0, 7.7, and 9.0 kb). This suggests multiple copies of genes encoding 16S rRNA.

Animals↗

PubMed2002

Identification of characteristic oligonucleotides in the bacterial 16S ribosomal RNA sequence dataset.

MOTIVATION: The phylogenetic structure of the bacterial world has been intensively studied by comparing sequences of 16S ribosomal RNA (16S rRNA). This database of sequences is now widely used to design probes for the detection of specific bacteria or groups of bacteria one at a time. The success of such methods reflects the fact that there are local sequence segments that are highly characteristic of particular organisms or groups of organisms. It is not clear, however, the extent to which such signature sequences exist in the 16S rRNA dataset. A better understanding of the numbers and distribution of highly informative oligonucleotide sequences may facilitate the design of hybridization arrays that can characterize the phylogenetic position of an unknown organism or serve as the basis for the development of novel approaches for use in bacterial identification. RESULTS: A computer-based algorithm that characterizes the extent to which any individual oligonucleotide sequence in 16S rRNA is characteristic of any particular bacterial grouping was developed. A measure of signature quality, Q(s), was formulated and subsequently calculated for every individual oligonucleotide sequence in the size range of 5-11 nucleotides and for 15mers with reference to each cluster and subcluster in a 929 organism representative phylogenetic tree. Subsequently, the perfect signature sequences were compared to the full set of 7322 sequences to see how common false positives were. The work completed here establishes beyond any doubt that highly characteristic oligonucleotides exist in the bacterial 16S rRNA sequence dataset in large numbers. Over 16,000 15mers were identified that might be useful as signatures. Signature oligonucleotides are available for over 80% of the nodes in the representative tree.

Algorithms↗

PubMed2026

Zhiling Jiangya decoction treats hypertension in rats: An integrative study of network pharmacology, immune infiltration, molecular simulation, and 16S rDNA sequencing.

OBJECTIVE: This study integrated network pharmacology, immune infiltration analysis, molecular docking, molecular dynamics simulation, ADMET prediction, 16S rDNA sequencing, and rat experiments to elucidate the potential mechanisms underlying the antihypertensive effects of Zhiling Jiangya Decoction (ZLJYD). METHODS: Active compounds and their potential targets were screened from the PubChem, TCMSP, NovoPro, and SwissTargetPrediction databases. Hypertension-related targets were retrieved from the OMIM and GeneCards databases, and overlapping targets were identified. The STRING database and Cytoscape 3.10.1 software were used to construct a protein-protein interaction network and a herb-component-target-disease network. Gene Ontology functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis were performed to identify the key biological processes and signaling pathways involved. Using the CIBERSORT algorithm combined with correlation analysis, we investigated the association between key targets and immune cell infiltration. Molecular docking, molecular dynamics simulations, and ADMET predictions were performed to assess the binding stability and pharmacokinetic properties of the main compounds with their corresponding targets. Finally, the antihypertensive efficacy of ZLJYD was validated using a spontaneously hypertensive rat model, and alterations in gut microbiota were analyzed using 16S rDNA sequencing. RESULTS: A total of 123 active compounds and 267 hypertension-related targets of ZLJYD were identified. Enrichment analysis revealed that these targets were primarily associated with the PI3K-Akt signaling pathway and lipid and atherosclerosis pathways. Immune infiltration analysis suggested that the therapeutic effects of ZLJYD may involve the regulation of follicular helper T cells, naïve B cells, and naïve CD4⁺ T cells. Molecular docking and dynamics simulations supported the stable binding of key compounds to their target proteins, while ADMET predictions indicated favorable pharmacokinetic properties and safety profiles. Rat experiments demonstrated that ZLJYD significantly reduced blood pressure in spontaneously hypertensive rats, partially alleviated gut microbiota dysbiosis, and altered microbial community structure and phylogenetic diversity. CONCLUSION: This study systematically elucidates the potential mechanisms underlying the antihypertensive effects of ZLJYD through multiple components, targets, and pathways, particularly immune regulation and gut microbiota remodeling. These findings provide mechanistic insights into its potential therapeutic application.

16S rDNA sequencing↗

PubMed1993

Rapid detection and identification of Mycobacterium avium by amplification of 16S rRNA sequences.

An assay that is based on the amplification of 16S rRNA sequences and that was initially developed to detect Mycobacterium paratuberculosis in cattle was used to test 20 serotypes of the Mycobacterium avium complex (MAC) and atypical mycobacterial species not belonging to MAC. Only serotypes 1 to 6 and 8 to 11, designated M. avium, were detected by the assay, indicating that it can be used for the rapid detection and identification of M. avium. The results of the assay for clinical samples from animals suspected of having mycobacterial infections indicated that it can also be used directly on clinical samples.

Animals↗

PubMed1995

An early origin of plastids within the cyanobacterial divergence is suggested by evolutionary trees based on complete 16S rRNA sequences.

It is generally accepted that the plastids arose from a cyanobacterial ancestor, but the exact phylogenetic relationships between cyanobacteria and plastids are still controversial. Most studies based on partial 16S rRNA sequences suggested a relatively late origin of plastids within the cyanobacterial divergence. In order to clarify the exact relationship and divergence order of cyanobacteria and plastids, we studied their phylogeny on the basis of nearly complete 16S rRNA gene sequences. The data set comprised 15 strains of cyanobacteria from different morphological groups, 1 prochlorophyte, and plastids belonging to 8 species of plants and 12 species of diverse algae. This set included three cyanobacterial sequences determined in this study. This is the most comprehensive set of complete cyanobacterial and plastidial 16S rRNA sequences used so far. Phylogenetic trees were constructed using neighbor joining and maximum parsimony, and the reliability of the tree topologies was tested by different methods. Our results suggest an early origin of plastids within the cyanobacterial divergence, preceded only by the divergence of two cyanobacterial genera, Gloeobacter and Pseudanabaena.

Biological Evolution↗